PROJECT SUMMARY Inflammatory bowel disease (IBD) is the result of exacerbated immune response against commensal or ?good? bacteria, whereas various gastrointestinal (GI) infections caused by ?bad? bacteria such as Salmonella can initiate the onset and relapse of IBD. How the protective immune response against ?bad? microbes is linked to the abnormal exacerbated immune response against ?good? microbes is unclear. Primary immune responses to GI infections occur in the context of broader secondary responses against commensals that breach damaged mucosa. Intestinal antigen-presenting cells (APCs) are heterogeneous immune cells that capture ?good? and ?bad? intestinal microbes and present them to T cells. T cells, after being educated by APCs, help to eradicate ?bad? microbes but become tolerant to ?good? microbes. Signals provided by APCs that include pro- and anti-inflammatory cytokines will determine whether T cells will recognize microbes as ?good? or as ?bad?. APC subsets responsible for presenting pathogenic and commensal bacteria to T cells are unknown. Macrophages (M?s) are the most numerous mucosal APCs but their role in adaptive immune responses against enteric pathogens has not been established. Our preliminary data show that mucosal M?s are heterogeneous; they induce protective immunity against Salmonella through coordinated efforts of three functionally distinct subsets by providing the innate immune control, initiating mucosal inflammation and activating T cells in the mesenteric lymph nodes (MLNs) where some M?s migrate upon infection. In this proposal, we will test the hypothesis that mucosal M?s, a driving force of protective immunity against Salmonella, play a central role in maintaining intestinal homeostasis after infection is cleared. We anticipate that post-infection, mucosal M?s re-establish the immunological tolerance to commensals through the balance between mucosa-resident and MLN-migratory M? subsets: 1) by switching their cytokine profile from pro- inflammatory to anti-inflammatory, and 2) by downregulating their migration to the MLNs to reduce interactions with T cells. Both processes are driven by sustained production of anti-inflammatory cytokines IL-10 and TGF?, and by reduced pro-inflammatory (TNF?) and Toll-like receptor (TLR) signaling in mucosal M?s following pathogen clearance, repair of the epithelial barrier and diminished translocation of commensal bacteria into the mucosa. Our hypothesis will be tested in mouse models of transient infectious and non-infectious colitis using mice depleted of mucosal M?s or Il10, Tgfb1, Ccr7, Tnf and Myd88 genes in M?s based on a Cre/loxP transgenic mouse approach. We anticipate that answering the questions raised in this proposal will provide new therapeutic strategies to reduce established inflammation and to prevent infection-driven IBD by promoting anti-inflammatory properties of mucosal M?s without compromising anti-microbial immunity.